11,15-dihydroxy-9-oxoprosta-8(12),13-dienoic acids and intermediate

ABSTRACT

The utilization of styrylglyoxal and a dialkyl ester of 3oxoundecane-1,11-dioic acid as starting materials leads to cisand trans-11,15-dihydroxy-9-oxoprosta-8(12),13-dienoic acids, which exhibit valuable pharmacological properties, e.g. hypotensive, antiulcerogenic and smooth muscle stimulating.

A [72] Inventor:

United States Patent Miyano [54] 11,15-DIHYDROXY-9-OXOPROSTA-8(l2),13-DIENOIC ACIDS AND INTERMEDIATE [63] Continuation-impart of Ser.No. 799,965, Feb.

17, 1969, abandoned.

[52] US. Cl .....260/514 R, 260/413, 260/520, 260/590, 424/317 [51] Int.Cl ..C07c 61/36 [5 8] Field of Search ..260/460, 714

Masateru Miyano, Morton Grove,

[4 1 Oct. 3, 1972 [56] References Cited UNITED STATES PATENTS 3,558,682l/ 1971 Pappo et a] ..260/468 Primary Exarniner-lnrraine A. WeinbergerAssistant Examiner-Robert Gerstl Attorney-John M. Brown, John J. Kolano,Elliot N. Schubert, Lowell C. Bergstedt, Sybil Meloy, Walter C. Ramm andl-lelmuth A, Wegner ABSTRACT The utilization of styrylglyoxal and adialkyl ester of 3- oxoundecane-l,l l-dioic acid as starting materialsleads to cisand trans-ll,l5-dihydroxy-9-oxoprosta- 8(l2),l3-dienoicacids, which exhibit valuable pharmacological properties, e.g.'hypotensive, antiulcerogenic and smooth muscle stimulating.

5 Claims, No Drawings l l l 5-DIl-IYDROXY-90XOPROSTA-8( l 2), l 3-DIENOIC ACIDS AND INTERMEDIATE Thisapplication is a continuation inpartof my copending application Ser. No. 799,965, filed Feb. 17, 1969, nowabandoned.

The present invention relates to novel prostanoic acid derivativescharacterized'by 8(12) and 13 double bonds and, more particularly, withl 1,15-dihydroxy-9- oxoprosta-8(l2),13-dienoic acids, as represented bythe following structural formula I COOH OH OH wherein the wavy linesindicate the alternative cis and trans relationship of the hydroxygroups.

A convenient procedure for manufacture of the compounds of thisinvention utilizes, as starting materials, styrylglyoxal, readilyprepared by the selenous acid oxidation of 4-phenyl-3-buten-2-one, and adialkyl ester of 3-oxoundecane-l,1ldioic acid. Dimethyl3-oxoundecane-l,l'l-dioate, for example, is saponified with potassiumhydroxide and the resulting dicarboxylic acid isallowed to react withstyrylglyoxal, thusaffording l4-phenyl-9, l 2-dioxo-l l-hydroxytetradecl3- enoic acid. Cyclizationof that acid by means of potassium hydroxideaffords 3-hydroxy-5-oxo-2-styrylcyclopent-l-eneheptanoic 'acid.Hydroxylation of the double bond with osmium tetroxide followed bycleavage of the resulting, glycol, typically with sodium periodate,affords 2-formyl-3-hydroxy-5-oxocyclopentl-eneheptanoic acid. Thataldehyde is allowed to react with hexanoylmethylene triphenylphosphorane to yield 1 l-hydroxy-9, l 5-dioxoprosta-8( l2),l 3-dienoicacid, which upon reduction, typically with sodium borohydride, affords amixture of the racemic cisl l,l5-dihydroxy and trans-l l,l5-dihydroxycompounds, i.e. ll,l5-dihydroxy-9'oxoprosta8(l2),l3- dienoic acid. Thesecis and trans racemates are separated chromatographically.

The compounds of this invention are obtained as racemic mixtures.Resolution of those racemates is conveniently achieved by standardmethods utilizing optically active amines such as the d or I enantiomersof brucine, morphine, quinine, quinidine, strychnine, menthylamine,cinchonine, cinchonidine and a-phenylethylamine. In that manner, trans-ll,l5-dihydroxy-9- oxoprosta-8(l2),l3-dienoic acid, for example, ,isresolved to afford its individual d and l enantiomers.

The compounds of this invention display valuable pharmacologicalproperties as is evidenced by their anti-ulcerogenic, hypotensive andsmooth muscle stimulating activity.

The anti-ulcerogenic property of the instant compounds is demonstratedby their ability to inhibit the ulceration reported by Shay et al.,Gastroenterology, 5, 43, (1945), to occur in rats subjected to fastingand pyloric ligation. In this test, male Charles River rats weighing200250 g. and fasted 72 hours prior to ligation are used. Immediatelyfollowing ligation, the prescribed dose of compound, dissolved orsuspended in [.0 ml. of pH 2.0 hydrochloric acid is intragastricallyadministered to each of a group of six animals. An initial dose of 50mg. per kg. of body weight is normally employed. A like group ofanimals'to which is identically and concurrently administered the acidalone serves as control. Precisely 19 hours later the stomachs ofsurviving animals are excised and examined under 5X magnification. Thenumber of ulcers occurring in a non-secretory portion of each stomach iscounted in four groups according to size, i.e. less than 2 mm., 2-4 mm.,4-8 mm. and greater than 8 mm. Each rat is then assigned a score, 2,which is aweighted average of the logarithms of the ulcer counts inseveral size groups determined by a formula found approximately optimalby discriminant function analysis to be as follows:

z= 20.00 log N, 1 +0.22 log (N2+ 1 +46.76 log (N3+ l)+6.ll log(N 1wherein N N, are the observed ulcer counts of the increasing sizegroups. Since long term studies in approximately 400 animals show thatthe average z value for controls is 96.2, with a standard error pergroup of six equal to 18.97, a decrease in the average z score for agiven test group, relative to concurrent controls, amounting to 37.5 ormore is significant (P 0.05); and a compound producing such a decreaseis considered anti-ulcerogenic.

For the determination of hypotensive activity the following assay isused:

Male Charles River rats weighing -350 g. are anesthetized byintraparitoneal injection of 1.5 g./kg. of urethane, whereuponcardiovascular reflexes are blocked by subcutaneous injection of 3 mg.of atropine sulfate dissolved in 0.3 ml. of aqueous 0.85 percent sodiumchloride and sensitization is induced by subcutaneous injection of 5 mg.of pentolinium tartrate dissolved in 1 ml. of aqueous 0.85 percentsodium chloride. The trachea is intubated and both femoral veins and acarotid artery are cannulated, the latter being connected to acalibrated transducer, amplifier and recorder. After surgery, 5 mg./kg.of heparin sodium is introduced via one of the venus cannulae as a 2percent solution in aqueous 0.85 percent sodium chloride and rectaltemperature is adjusted to 35 C. by means of a regulator and externalheat source. When the animals blood pressure and temperature havestabilized, a high and a low dose, with a ratio of high dose to low doseof 2:1, of the test compound and a high and a low dose, also in theratio of 2:1, of the standard prostaglandin B are individuallyadministered intravenously and in a random pattern and the bloodpressure measured after each injection. The blood pressure of eachanimal is permitted to return to normal between successive injections.The relative potency of the test compound compared to the standardprostaglandin E is determined by a four point bioassay using arandomized block design. The results are analyzed by the method of C. I.Bliss, The Statistics of Bioassay,-,.Academic Press, New York 1952).

The smooth muscle stimulating property of the compounds of thisinvention is demonstrated by their activity in each of the followingthree assays:

A segment of one of the uterine horns of a freshly killed rat is removedafter determining that the rat is in the diestrus phase of the ovariancycle by microscopic examination of a vaginal washing. The segment ismountedin a 2 ml. tissue bath containing de .Ialon solution maintainedat 37 C. and bubbled with a gaseous mixture of 95 percent oxygen andpercent carbon dioxide. Longitudinal contractions of the tissue elicitedby l mcg./ml. and 2 mcg./ml. of the test compound are measuredisotonically and compared to those elicited by 0.1 mcg./ml. and 0.2mcg./ml. ofa standard preparation of prostaglandin E The relativepotency of the test compound compared to the standard prostaglandinpreparation is determined by a four point bioassay using a randomizedblock design. Results are analyzed by the aforementioned method ofBliss.

In the second assay, a segment of duodenum from a freshly killed rabbitis mounted in a 4 ml. tissue bath containing Tyrode solution maintainedat 37 C., through which is bubbled a gaseous mixture of 95 percentoxygen and 5 percent carbon dioxide. Longitudinal contractions of thetissue educed by 0.625 mcg./ml. and 1.25 mcg./ml. of the test compoundare measured isotonically and compared to those educed by 0.025 meg/ml.and 0.05 mcg./ml. of a standard preparation of prostaglandin E Therelative potency of the test compound compared to that of the standardis determined by a four point bioassay using a randomized block designand results are analyzed by the aforementioned method of Bliss.

In the third assay, a segment of distal ileum from a freshly killedguinea pig is mounted in a 2 ml. tissue bath containing modified Tyrodesolution with one-half of the usual concentration of magnesium ions. Thetemperature is maintained at 37 C. and the solution is bubbled with agaseous mixture of 95 percent oxygen and 5 percent carbon dioxide.Longitudinal contraction of the tissue elicited by 5 mcg./ml. andmcg./ml. of the test compound are measured isotonically and compared tothose elicited by 0.05 mcg./ml. and 0.1 meg/ml. of a standardpreparation of prostaglandin E The relative potency of the test compoundis determined by the procedure of Bliss described above.

Catalytic hydrogenation of the instant compounds results in saturationof the 8(12) and 13 olefinic linkages, thus affording the prostanoicacid derivatives of the following formula wherein the wavy linesindicate the alternative cis and trans relationship of the hydroxygroups. As a specific example, trans-l l l 5-dihydroxy-9-oxoprosta-812),1 3-dienoic acid is hydrogenated in the presence of a 5 percentpalladium-on-carbon catalyst to afford trans-ll,l5-dihydroxy-9-oxoprostanoic acid. The compounds encompassed by thelatter formula similarly are valuable pharmacological agents. Theypossess, for example, hypotensive activity, as is demonstrated in theassay described hereinbefore.

The invention will appear more fully from the examples which follow.These examples are given by way of illustration only and are not to beconstrued as limiting the invention either in spirit or in scope sinceit will be apparent to those skilled in the art that many modificationsboth in materials and methods may be practiced without departing fromthe purpose or intent of this disclosure. Throughout these examplestemperatures are given in degrees Centigrade (C.) and relative amountsof materials in parts by weight except as otherwise noted.

EXAMPLE 1 A solution containing 100 parts of 4-phenyl-3-buten- 2-one,106 parts of selenous acid, 160 parts of dioxane and 20 parts of wateris heated to the reflux temperature. After the initial vigorous reactionhas subsided, the mixture is heated at that temperature for anadditional 30 minutes. The supernatant is then decanted from themetallic selenium and is concentrated under reduced pressure.Distillation of the residue under reduced pressure affordsstyrylglyoxal, boiling at about 120 at 2.5 mm. pressure.

EXAMPLE 2 A solution of 38.2 parts of dimethyl 3-oxoundecanel,l l-dioatein 200 parts by volume of 10 percent aqueous potassium hydroxide isstored at O-5 for about 3 days, then is adjusted to pH 5 by the additionof concentrated aqueous citric acid. To that mixture is added a solutionwhich is prepared by heating 21.9 parts of styrylgyloxal in 50 parts byvolume of 50 percent aqueous methanol at 6575 for about 20 minutes, thenadding 60 parts of methanol. To the resulting reaction mixture is added30 parts by volume of l M pH 4.5-5.0 citrate buffer and stirring at roomtemperature is continued for about 3 hours, during which time carbondioxide gas is evolved. The precipitated product is collected byfiltration, thus affording the half potassium salt of l4-phenyl-9, 12-dioxo-l l-hydroxytetradecl 3- enoic acid, melting at about 105.Further purification by recrystallization from methanol affords the purecompound, melting at about l07.5.

The latter half potassium salt is dissolved in water and the resultingaqueous solution is acidified by the addition of dilute hydrochloricacid. The resulting acidic mixture is extracted with ether and the etherlayer is separated, washed with water, dried over anhydrous sodiumsulfate and concentrated to dryness. The resulting solid residue ispurified by recrystallization from chloroform-ether to yieldl4-phenyl-9,l2- dioxo-l l-hydroxytetradec-l3-enoic acid, melting atabout 8 l .5-83.

EXAMPLE 3 To 3,000 parts by volume of an aqueous solution containing 6.7parts of potassium hydroxide is added, with stirring at about 2l23 overa period of about 2 hours, a solution of 10.4 parts of l4-phenyl-9,l2-dioxo-l l-hydroxytetradec-l3-enoic acid in 187 parts of chloroform.After completion of the addition, the reaction mixture is stirred for anadditional 2 hours, then is made acidic by adding 10 parts of oxalicacid dihydrate. The acidic mixture is extracted with chloroform and theorganic layer is washed with dilute aqueous sodium chloride, then driedover anhydrous sodium sulfate and concentrated to dryness under reducedpressure. The resulting residue is recrystallized first from benzene,then from chloroform-ether to yield 3-hydroxy-5-oxo-2-styrylcyclopentl-eneheptanoic acid, which displays a melting point at about 1 18. Thiscompound exhibits an ultraviolet absorption maximum at about 325millimicrons with a molecular extinction coefficient of about 36,400.

EXAMPLE 4 A mixture consisting of 13 parts of 3-hydroxy-5-oxo-2-styrylcyclopent-1-eneheptanoic acid, 17.8 parts of sodium periodate,55 parts of water, 160 parts of dioxane and 2 parts of a 2 percentosmium tetroxide in dioxane solution is stirred under nitrogen at roomtemperature for about 4 hours. The reaction mixture is then extractedwith ether and the ether layer is separated and extracted several timeswith 0.5 percent aqueous sodium chloride. The salt extracts aresaturated with sodium chloride, then extracted with ether. The etherlayer is separated, dried over anhydrous sodium sulfate, thenconcentrated under reduced pressure to afford2-formyl-3-hydroxy-5-oxocyclopent-1- eneheptanoic acid, characterized byan ultraviolet absorption maximum at about 228 millimicrons with amolecular extinction coefficient of about 10,100.

EXAMPLE 5 To a solution of 10.2 parts of 2-formyl-3-hydroxy-5-oxocyclopent-l-eneheptanoic acid in 200 parts of dioxane is added 4parts of triethylamine and the resulting mixture is stripped of excesstriethylamine by distillation under reduced pressure. The resultingresidue is dissolved in 210 parts of dioxane. To that dioxane solutionis then added 15.3 parts of hexanoylmethylene triphenyl phosphoranedissolved in 396 parts of benzene. The resulting reaction mixture isheated at the reflux temperatureunder nitrogen for about 18 hours, thenis cooled, washed with aqueous oxalic acid and extracted with aqueouspotassium bicarbonate. That alkaline extract is washed with ether, thenacidified with oxalic acid and extracted with ether. The ether extractis washed with aqueous sodium chloride, dried over anhydrous sodiumsulfate and concentrated to dryness under reduced pressure to afford thecrude product, which is purified by dry column chromatography on silicagel containing 8 percent of water, using 4 percent methanol in benzene,or by chromatography on silica gel and elution with 40 percent ethylacetate in benzene, thus affording l l-hydroxy-9,l5-dioxoprosta- 8(12),13-dienoic acid, characterized by an ultraviolet absorption maximum atabout 291 millimicrons with a molecular extinction coefficient of about21,900.

EXAMPLE 6 To a solution of 12 parts ofll-hydroxy-9,l5-dioxoprosta-8(l2),l3'dienoic acid in 28 parts ofethanol, cooled to 0-5, is added dropwise a solution of 3 parts oftriethylamine in 275 parts of water. To that mixture is added dropwisewith cooling and stirring a solution of 0.32 part of sodium borohydridein 32 parts of water. Stirring at approximately 10 is continued forabout 25 minutes, at the end of which time the reaction mixture ispoured carefully into excess aqueous citric acid. Extraction with etheraffords an organic solution, which is washed with water, dried overanhydrous sodium sulfate and concentrated under reduced pressure toafford l l l 5-dihydroxy-9-oxoprosta-8( 12), l 3-dienoic acid. Thismixture of epimeric ll-hydroxy compounds is separated by chromatographyon silica gel, using 25-50 percent ethyl acetate in benzene mixtures asthe eluant. The 40 percent ethyl acetate in benzene eluant yieldssuccessively cis-l 1 ,15-dihydroxy-9-oxoprosta- 8, l 2( l3 )-dienoicacid and trans-1 1,15-dihydroxy-9-oxoprosta-8,l2(l3)-dienoic acid.Further purification is achieved by partition chromatography on silicicacid, wherein the solvents are prepared by shaking together 500 parts byvolume of methanol, 200 parts by volume of distilled water and 1,500parts by volume of benzene. The lower layer is used as the stationaryphase and the upper layer as the eluant. Successive elutions of thecolumn with that eluant affords first trans-11,15,-dihydroxy-9-oxoprosta-8( l2),l 3-dienoic acid, which exhibits anultraviolet absorption maximum at about 276 millimicrons with amolecular extinction coefficient-of about 26,500, thencis-l1,15-dihydroxy-9-oxoprosta-8(12),13-dienoic acid, exhibiting anultraviolet absorption maximum at about 276 millimicrons with amolecular extinction coefficient of about 27,800.

EXAMPLE 7 To a solution of 0.96 part of cis-ll,15-dihydroxy-9-oxoprosta-8( l2),l3-dienoic acid in 40 parts of isopropyl alcohol isadded 0.2 part of 5 percent palladium-on-carbon catalyst and theresulting mixture is shaken with hydrogen under 40-52 pounds per squareinch pressure at about 53 for approximately 48 hours. The catalyst isthen removed by filtration and the filtrate is concentrated to drynessunder reduced pressure. The resulting residue is dissolved in 480 partsof percent ethanol containing 30 parts of potassium acetate and thatmixture is kept at room temperature for about 95 hours. At the end ofthat time the solvent is removed by distillation'under reduced pressureand the resulting residue is dissolved in approximately 400 parts ofwater. That aqueous solution is acidified with citric acid to pH 3.5,then is extracted with ether. That organic solution is washed withdilute aqueous sodium chloride, then dried over anhydrous sodium sulfateand concentrated under reduced pressure. The resulting crude product ispurified by partition chromatography on silicic acid, wherein thesolvents are prepared by shaking together 1,500 parts by volume ofbenzene, 500 parts by volume of methanol and 200 parts by volume ofwater. The lower layer is used as the stationary phase and the upperlayer as the eluant. Elution of the column in that manner affordssuccessively d1- l1B,l5(R)-dihydroxy-9-oxoprostanoic acid and d1- 11a,15(S)-dihydroxy-9-oxoprostanoic acid.

EXAMPLE 8 A mixture containing 1.16 parts of trans-11,15-dihydroxy-9-oxoprosta-8(12),13-dienoic acid, 40 parts of isopropylalcohol and 0.2 part of 5 percent palladium-on-carbon catalyst is shakenwith hydrogen under 38-52 pounds per square inch pressure at about 50for approximately 48 hours, then is concentrated to dryness underreduced pressure. The resulting residue is contacted with a solution ofpotassium acetate in 95 percent ethanol according to the proceduredescribed in Example 7, and the product is isolated from that mixture bythe procedure described in that Example. Purification of that crudeproduct by partition chromatog- 7 8 raphy in the manner described inExample 7 results in, 2. As in claim 1, the compound which is 11,15- ucci y 1 l (S ihydrXy-9-oxoprostanoic dihydroxy-9-oxoprosta-8( l2), 13-dienoic acid. mild and ('1111,15(R)'dlhydroxy-9'oxoprostanoic 3. As inclaim 1, the compound which is cis-l 1,15-

acid.

What is claimed is: l. A compound of the formuladihydroxy-9-oxoprosta-8( l2), 1 3-dienoic acid. 5 4. As in claim 1, thecompound which is trans-l 1,15-

dihydroxy-9-oxoprosta-8( l 2),1 3-dienoic acid.

0 5. 1 l-l-Iydroxy-9,15-dioxoprosta-8( l2),13-dienoic l c 0 0H acid

2. As in claim 1, the compound which is11,15-dihydroxy-9-oxoprosta-8(12),13-dienoic acid.
 3. As in claim 1, thecompound which is cis-11,15-dihydroxy-9-oxoprosta-8(12),13-dienoic acid.4. As in claim 1, the compound which istrans-11,15-dihydroxy-9-oxoprosta-8(12),13-dienoic acid. 5.11-Hydroxy-9,15-dioxoprosta-8(12),13-dienoic acid.